Autosomal dominant polycystic kidney disease (ADPKD) accounts for approximately 2% of end-stage renal disease. Recent advances in characterization of the PKD1 and PKD2 genes, particularly through cell culture studies, have suggested multiple potential mechanisms for cyst formation. What is needed now is an ADPKD model that can be easily manipulated to test the importance of these pathways. The zebrafish may bridge the gap between the limits of in vitro studies and the difficulties inherent to studying rodent models. The sequencing of the zebrafish genome and the ease of performing genetic manipulations through embryo microinjection have made this organism a powerful tool for studying vertebrate genetics. We have cloned a zebrafish ortholog to PKD2. Blockade of its expression produces bilateral renal cysts. In Aim 1, we will perform an in vivo structure-function analysis of ADPKD gene products via transient knock-in/knock-down techniques to examine (a) the functional role of well-described in vitro PKD2 interactions and (b) the cystpromoting activity of the PKD1 cytoplasmic domain. In Aim 2, we will assess the contributions of two downstream pathways to cyst formation combining genetic and pharmacologic methods: cAMP/EGFR signaling and angiogenesis. We hope to capitalize on this high through-put model to elucidate in vivo functions of ADPKD gene products, identify novel in vivo targets that modify cyst formation, and show that the zebrafish is a useful pre-rodent drug-screening tool for this important disease. This work will be performed at Harvard Medical School and Beth Israel Deaconess Medical Center, in the laboratory of Vikas P. Sukhatme, M.D., Ph.D. New skills the applicant will learn include development of a disease model, structure-function analysis, and signal transduction work in vivo. The applicant has spent the last year generating the data presented herein and expects to publish papers shortly on zebrafish models of cystic and proteinuric diseases. He is committed to a career centered on basic research in an academic renal division. The expertise of the mentor and the environment of Harvard Medical School--along with the applicant's prior research, the complexity of the proposed project, and his ongoing commitment to learn and apply molecular biology to understand human disease-provide a unique opportunity for the applicant to achieve the goals of the K award and launch an independent research career.